Oligodendrocytes (OLs) are myelin-forming cells in the central nervous system (CNS). CNS myelin formation/repair consists of two closely-related sequential events: OL differentiation from oligodendrocyte progenitor cells and axonal (re)myelination by already differentiated OLs. Defects of these two events result in abnormalities of CNS myelin formation such as in periventricular leukomalacia and inability of myelin repair such as in multiple sclerosis. Our long term goal is to study the underlying mechanisms regulating CNS myelin formation/repair. The expression of the Wnt effector transcription factor 7-like 2 (TCF7l2, a.k.a. TCF4) in multiple sclerosis lesions is one such promising mechanism. A well-studied role of TCF7l2 is transcriptionally mediating Wnt/?-catenin signaling in Wnt activated cells such as colorectal cancer cells. Previous studies from others and our own laboratory have shown that genetic activation of canonical Wnt/?-catenin signaling pathway inhibits OL differentiation (review Guo et al., 2015).Therefore, it has been proposed that TCF7l2 inhibits OL differentiation acting through Wnt/?-catenin signaling. However, we recently reported that conditionally disrupting TCF7l2 by Cre-loxP genetic approach inhibits neonatal and early postnatal OL differentiation without perturbing Wnt/?-catenin signaling pathway (Hammond et al., 2015). Based on our genetic data, we propose an alternative hypothesis that TCF7l2, acting through non-Wnt pathways (Aim 2), is a multimodal positive regulator of CNS myelin formation (Aim 1) that can be manipulated to promote CNS myelin repair after myelin damage (Aim 3). In Aim 1, we will specifically ablate TCF7l2 in already differentiated OLs to determine its role in subsequent axonal myelination and myelin lipid synthesis. The experiments in this Aim will reveal a previously unrecognized novel role of TCF7l2 in CNS myelination independent of upstream OL differentiation. In Aim 2, we will use in vivo and in vitro genetic approaches to test the hypothesis that TCF7l2's function as a gene repressor plays an essential role in CNS myelin formation. This project will unveil a novel link between TCF7l2 and an oligodendroglial autocrine pathway that inhibits OL differentiation and myelination. Our paradigm- shifting data make it necessary and important to revisit and reevaluate the therapeutic potential of TCF7l2 during CNS myelin repair which has been proposed as an inhibitory factor in human multiple sclerosis. In Aim 3, we will use lentiviral vector-mediated gene transfer to test the alternative hypothesis that enforced TCF7l2 expression promotes OL differentiation and CNS myelination in demyelination animal models. The expected overall impact of this innovative proposal is that it will fundamentally advance our mechanistic understanding of CNS myelin formation and of novel role of TCF7l2, and will change our conventional view of inhibiting TCF7l2 to enhance CNS myelin repair.